DNA Profiling of Transgenes in Genetically Modified Plants
Bittsánszky A
1,2
, Gabor G
1*
Gullner G and Komíves T
1
Plant Protection Institute, CAR, Hungarian Academy of Sciences, Hungary
2
Inst Genetics and Biotechnology, Szent István University, Hungary
*
Corresponding author: Gyulai Gabor, Inst Genetics and Biotechnology, Szent István University, 2103 Gödöllő, Páter K. u. 1.Godollo, Hungary, Tel: +36 28 522-069; E-
mail: gyulai.gabor@mkk.szie.hu
Received date: April 20, 2016; Accepted date: May 25, 2016; Published date: June 02, 2016
Copyright: © 2016 Bittsánszky A, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution and reproduction in any medium, provided the original author and source are credited.
Abstract
In silico sequence diversities of four orthologous plant gsh1 genes and their anino acid translates of GSH1
proteins (Glutathione Synthase) were compared to the non-orthologous prokaryotic gshI/GSHI gene/protein of E. coli
(NCBI # X03954). Primer pair was designed and transgene detection was carried out in two types of gshI-transgenic
poplar clones (Populus x canescens) of ggs11 (cyt-ECS) and lgl6 (chl-ECS). Usefulness of genetic modification
technologies (GMO) is indicated.
Keywords: Biotechnology; DNA Proiling
Introduction
Transgenes represent genetic markers artiicially introduced in
laboratory motivated to improve crops. Detection of marker in the
Genetically Modiied Organism (GMO), and its vegetative or sexual
progenies; and monitoring it in test and cultivated populations as well
as in exposed non-target cross-pollinated populations is of
fundamental and practical importance. he genetically modiied state
of an organism, i.e. the presence of the transgene, is veriied essentially
by DNA proiling. Selecting the DNA sequence for DNA proile is
straightforward because a known sequence is introduced. Introduction
of genes, self or foreign, into plants had prerequisites. he ability to
select and identify desired genotypes in cells, tissues or intact plants
laid the fundamentals for application of genetic transformation of
plants and animals by tools of biotechnology.
he Biological Research Centre (Szeged, Hungary) can be
considered as the Genius Loci [1] of the current plant biotechnology
since methodologies of plant cell line selections for chloroplast
mutants [2,3], cell fusion [4], genetic transformation [5,6] bacterial
nitrogen ixation [7] and artiicial chromosomes [8] were either
fundamentally developed or highly improved there.
he irst stable higher plant mutant, the antibiotic (i.e. streptomycin,
SR) resistant (i.e. mutant) tobacco (SR1) was selected [2] in vitro,
followed by the selection [9] and identiication of SR1A15 [10] the irst
double mutant of higher plants, the albino (chloroplast) tobacco [9,11].
Later, as the early forms of gene transfer, protoplast cell fusion
plants (i.e. cybrids) were developed in several laboratories [12-16].
Alternatives to the conventional haploid genome transfer (i.e,
pollination), the technologies of single and pyramided gene transfer
resulting in stable transgenic crops (i.e. GM - genetically modiied, or
GMO - genetically modiied organism), were developed in four labs at
the same time in 1983: GM Nicotiana plumbaginifolia (resistant to the
antibiotic kanamycin) [17], other tobacco lines resistant to kanamycin
and methotrexate (a drug used to treat cancer and rheumatoid
arthritis) [18], GM petunia resistant to kanamycin [19] and GM
sunlower transformed by phaseolin gene isolated from bean [20].
he irst ield trial of GM cotton was carried out in 1990, followed
by the irst FDA-approved (Unites States Food and Drug
Administration) transgenic food of Flavr-Savr tomato in 1994 [21]. A
series of further GM crops were released in 1995, such as the canola oil
seed rape (Brassica napus) with modiied oil composition (Calgene), Bt
(Bacillus thuringiensis) corn (Ciba-Geigy) resistant to the herbicide
bromoxynil (Calgene), Bt cotton (Monsanto), GM soybeans resistant
to herbicide glyphosate (Monsanto); virus-resistant squash (Asgrow),
and a delayed ripening tomatoes (DNAP, Zeneca/Peto and Monsanto)
[22,23]. Later, a series of woody plants were also bred by genetic
transformation [24-29].
Here we present a case study of barcoding (i.e. detecting and
monitoring GM plants) the CaMV-35S-gshI poplar (Populus x
canescens) with techniques useful for both developing GM plants and
for anti-GM purposes.
Materials and Methods
DNA extraction
Total DNA samples of 0.1 g leaf tissue in each case were extracted in
CTAB, cethyltrimethylammonium bromide, bufer followed by RNase-
A (from bovine pancreas, Sigma, R-4875, treatment) for 30 min at 37C.
DNA samples of ten individuals of each line were pooled in one bulk
and subjected to PCR analysis.
Multiple sequence alignments for primer design
Nucleotide sequences of genes gsh1 were downloaded from the
National Center for Biotechnology Information (NCBI) databases [30].
Multiple sequence alignments were applied in silico with the sotware
programs BioEdit Sequence Alignment Editor (North Carolina State
University, USA) [31], Multalin [32], Clustal W [33], FastPCR [34] and
computer program MEGA4 [35].
Forensic Biomechanics
Gabor et al., J Forensic Biomed 2016, 7:2
http://dx.doi.org/10.4172/2090-2697.1000129
Research Article Open Access
J Forensic Biomed
ISSN:2090-2697 JFB, an open access journal
Volume 7 • Issue 2 • 1000129